Proteomic profiling of pseudorabies virus-infected PK-15 cells based on 4D label free analysis

Abstract

Pseudorabies virus (PRV) heavily depends on host machinery to support its life cycle. Investigating the interaction between PRV and host could aid in the understandings of viral pathogenesis. In this study, we performed a 4D label free proteomic method to examine the differentially expressed proteins in porcine kidney PK-15 cells with PRV infection. The results showed that the levels of 661 proteins were significantly elevated and 693 proteins were markedly reduced. Furthermore, these altered proteins were primarily enriched in spliceosome, protein processing in endoplasmic reticulum (ER), RNA transport, and protein export. To ensure the reliability of the proteomic results, the protein levels of formin binding protein 11 and wolfram syndrome 1as components of spliceosome and ER were verified via western blotting and the results were consistent. Together, our data shed light on a new protein profiling induced by PRV infection and highlighted the importance of spliceosome and ER in PRV replication which could promote understandings of host-PRV interplay.

Keywords: Endoplasmic reticulum; Herpes virus; Proteomic analysis; Spliceosome.

Fig. 1

Overview of the study. A) Principal component analysis. B) The number of mass distribution. C) The number of peptides per protein. D) The number of detected peptides and proteins. PRV: Pseudorabies virus

Fig. 2

Analysis of differentially expressed proteins. A) The volcano plots showed the downregulated proteins (Green), the upregulated proteins (Orange), and the unchanged proteins (Grey). B) Statistical analysis of changed proteins. C) The heat map showed the changes of proteins. D) Analysis of Clusters of Orthologous Groups (COG)/euKaryotic Orthologous Groups (KOG) pathways.

Fig. 3

The subcellular location illustrated the distribution of altered proteins.

Fig. 4

Analysis of the enriched pathways. A) Biological process, B) Cellular component, C) Molecular function, and D) Proteins' Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment.

Fig. 5

The network in spliceosome and endoplasmic reticulum. A) The composition of spliceosome, B) Protein processing in endoplasmic reticulum. The proteins in green were reduced, while the proteins in red were increased.

Fig. 6

A) PK-15 cells were infected with pseudorabies virus (PRV; multiplicity of infection = 0.50, 1.00, and 2.00) or mock-infected with Dulbecco's modified Eagle's medium for 40 hr. Cells were lysed and probed for the levels of formin binding protein 11 (FBP11) and tubulin. B) The levels of wolfram syndrome 1 (WFS1) and tubulin were examined from the same samples by western blotting.